Composite Microneedle Array Including Nanostructures Thereon

ABSTRACT

Disclosed are composite microneedles arrays including microneedles and a film overlaying the microneedles. The film includes a plurality of nano-sized structures fabricated thereon. Devices may be utilized for interacting with a component of the dermal connective tissue. A random or non-random pattern of structures may be fabricated such as a complex pattern including structures of differing sizes and/or shapes. Devices may be beneficially utilized for delivery of an agent to a cell or tissue. Devices may be utilized to directly or indirectly alter cell behavior through the interaction of a fabricated nanotopography with the plasma membrane of a cell and/or with an extracellular matrix component.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims priority to U.S. Provisional Patent ApplicationSer. No. 61/328,723 having a filing date of Apr. 28, 2010, U.S.Provisional Patent Application Ser. No. 61/411,071 having a filing dateof Nov. 8, 2010, and United States Provisional Patent Application Ser.No. 61/435,939 having a filing date of Jan. 25, 2011, all of which areincorporated herein in their entirety by reference.

BACKGROUND

Primary drug delivery methods include oral delivery and injections, butthese methods present difficulties. For instance, injections are painfuland both methods tend to provide bursts of agents rather than apreferred steady-state delivery. Additionally, the successful long termuse of both oral delivery and injected delivery requires the patient toconsistently meet the time requirements for the delivery method.

Transdermal delivery materials have been developed in an attempt toprovide a painless route for delivery of active agents over a sustainedperiod with little or no interruption of the patient's daily routine.Unfortunately, natural dermal characteristics such as the overlappingcorneocytes of the stratum corneum, the tight junction of the stratumgranulosum, and Langerhans cells of the stratum spinosum that mayinstitute an immune response and/or a foreign body response all presentbarriers to successful transdermal delivery of an active agent.

Devices including microneedles that may facilitate transdermal deliveryof active agents have improved transdermal delivery. A microneedletransdermal device includes an array of needles that may penetrate atleast the stratum corneum of the skin and reach an underlying layer ofthe skin. In some devices, the microneedles are designed so as topenetrate to a depth that does not stimulate the nerve endings andinstitute a pain response. Examples of microneedle devices have beendescribed in U.S. Pat. No. 6,334,856 to Allen, et al. and U.S. Pat. No.7,226,439 to Prausnitz, et al., both of which are incorporated herein byreference.

Unfortunately, even with the inclusion of microneedles on a transdermaldevice, transdermal devices are presently limited to delivery of lowmolecular weight agents that have a moderate lipophilicity and nocharge. Even upon successful crossing of the natural dermal boundary,problems still exist with regard to maintaining the activity level ofdelivered agents and avoidance of foreign body and immune response.

The nanotopography of a surface adjacent to a cell has been found toaffect adhesive characteristics between the two as well as to effectcell behavior including morphology, motility, cytoskeleton architecture,proliferation, and differentiation (see, e.g., Hart, et al., EuropeanCells and Materials, Vol. 10, Suppl, 2, 2005; Lim, et al., J R SocInterface, Mar. 22, 2005, 2(2), 97-108; Yim. et al., Biomaterials, Sep.,2005, 26(26), 5405-5413). As an extension of this initial research,nanotopography of supporting substrates has been examined for use intissue engineering (see, e.g., U.S. Patent Application Publication Nos.2008/0026464 to Borenstein, et al, and 2008/0311172 to Schapira, etal.).

What are needed in the art are improved drug delivery devices. Forinstance, devices that provide efficient delivery of active agents whiledecreasing potential immune and foreign body response to both thedelivery device and the delivered agents would be beneficial.

SUMMARY

According to one embodiment, disclosed is a composite microneedle array.An array may include a microneedle assembly that includes a supporthaving a first surface and a second surface, wherein a plurality ofmicroneedles extend outwardly from the first surface. An array may alsoinclude a film overlaying the microneedles of the microneedle assemblythat at least partially conforms to the shape of the microneedles. Thefilm may have a first surface and a second surface. The first surface ofthe film may be adhered to the microneedle assembly, and the secondsurface of the film may include thereon a plurality of nanostructures,the nanostructures being arranged in a predetermined pattern.

Also disclosed is a method for forming a composite microneedle array. Amethod may include laying a film over a microneedle assembly andengaging the film with the microneedle assembly such that the film atleast partially conforms to the microneedles of the array and adheres tothe microneedle assembly.

BRIEF DESCRIPTION OF THE DRAWINGS

A full and enabling disclosure of the subject matter, including the bestmode thereof, directed to one of ordinary skill in the art, is set forthmore particularly in the remainder of the specification, which makesreference to the appended figures in which:

FIG. 1 schematically illustrates one embodiment of a microneedle array.

FIG. 2 is a scanning electron micrograph (SEM) image of anotherembodiment of a microneedle array,

FIGS. 3 and 4 are partial cross-sectional views of microneedle arraysthat may be formed in accordance with an embodiment of the presentdisclosure;

FIGS. 5A and 5B illustrate opposite sides of a nanopatterned film as maybe applied to a microneedle array.

FIG. 6 illustrates one embodiment of a complex pattern that may beformed on a film surface.

FIG. 7 illustrates a pattern including multiple iterations of thecomplex pattern of FIG. 6.

FIGS. 8A-8D illustrate examples of complex fractal and fractal-likenanotopographies.

FIG. 9 illustrates another complex pattern that may be formed on a filmsurface.

FIGS. 10A-10C illustrates exemplary packing densities as may be utilizedfor nano-sized structures as described herein including a square packingdesign (FIG. 10A), a hexagonal packing design (FIG. 10B), and a circlepacking design (FIG. 10C).

FIG. 11 illustrates the fractal pattern known as the Sierpenskitriangle.

FIG. 12 schematically illustrates one embodiment of a microneedleincluding a film at a surface that defines a nanotopography that mayinteract with an extracellular matrix (ECM).

FIG. 13 schematically illustrates a method of draping a nanopatternedfilm over a microneedle of an array to form a composite microneedlearray,

FIGS. 14A and 14B schematically illustrate one embodiment of a drugdelivery device in an exploded view (FIG. 14A) and assembled (FIG. 14B),

FIG. 15 is a perspective view of one embodiment of a transdermal patchprior to delivery of a drug compound.

FIG. 16 is a front view of the patch of FIG. 15.

FIG. 17 is a perspective view of the patch of FIG. 15 in which therelease member is partially withdrawn from the patch.

FIG. 18 is a front view of the patch of FIG. 15.

FIG. 19 is a perspective view of the transdermal patch of FIG. 15 afterremoval of the release member and during use.

FIG. 20 is a front view of the patch of FIG. 19.

FIG. 21 is a perspective view of another embodiment of a transdermalpatch prior to delivery of a drug compound.

FIG. 22 is a front view of the patch of FIG. 21.

FIG. 23 is a perspective view of the patch of FIG. 21 in which therelease member is partially peeled away from the patch.

FIG. 24 is a front view of the patch of FIG. 23.

FIG. 25 is a perspective view of the patch of FIG. 21 in which therelease member is completely peeled away from the patch.

FIG. 26 is a perspective view of the transdermal patch of FIG. 21 afterremoval of the release member and during use.

FIGS. 27A-27E illustrate several nanotopography patterns as describedherein.

FIG. 28 is an SEM of a film including a nanopatterned surface.

FIGS. 29A and 29B are two SEM of a film including another nanopatternedsurface.

FIG. 30 is an SEM of a film including another nanopatterned surface.

FIG. 31 is an SEM of a film including another nanopatterned surface.

FIG. 32 is an SEM of a film including another nanopatterned surface.

FIG. 33 is an SEM of a film including another nanopatterned surface.

FIG. 34 is an SEM of a film including another nanopatterned surface.

FIG. 35 is an SEM of a film including another nanopatterned surface.

FIG. 36 is an SEM of a film including another nanopatterned surface.

FIG. 37 is an array of microneedles including a film draped on themicroneedles, the film defining a pattern of nanostructures thereon.

FIG. 38 is a single microneedle of the array of FIG. 37.

FIG. 39 graphically illustrates the PK profile of a protein therapeuticdelivered with a device as illustrated in FIG. 37.

FIG. 40A and 40B are cross sectional images of skin followingtransdermal delivery of a protein therapeutic across the skin. FIG. 40Ais a cross section of skin that was in contact with a transdermal devicedefining nanotopography thereon, and FIG. 40B is a cross section of skinthat was in contact with a transdermal device including no pattern ofnanotopography formed thereon.

FIG. 41 graphically illustrates the blood serum concentration of aprotein therapeutic delivered with a device as described herein.

DETAILED DESCRIPTION OF REPRESENTATIVE EMBODIMENTS

Reference now will be made in detail to various embodiments of thedisclosed subject matter, one or more examples of which are set forthbelow. Each example is provided by way of explanation, not limitation.In fact, it will be apparent to those skilled in the art that variousmodifications and variations may be made in the present disclosurewithout departing from the scope or spirit of the subject matter. Forinstance, features illustrated or described as part of one embodimentmay be used on another embodiment to yield a still further embodiment.Thus, it is intended that the present disclosure covers suchmodifications and variations as come within the scope of the appendedclaims and their equivalents.

In general, a microneedle array is disclosed herein. The microneedlearray is a composite structure that includes a microneedle assembly anda film applied to the surface of the assembly and contacting themicroneedles. The film defines a plurality of fabricated structures on asurface. At least a portion of the structures are fabricated on ananometer scale. As utilized herein, the term ‘fabricated’ generallyrefers to a structure that has been specifically designed, engineered,and/or constructed so as to exist at a surface and is not to be equatedwith a surface feature that is merely an incidental product of aformation process. Thus, there will be a predetermined pattern ofnanostructures on the surface of the microneedles.

Also disclosed is a method of forming the composite arrays. A formationprocess may include the draping or layering of the film over themicroneedle assembly such that the film at least partially conforms tothe shape of the microneedles and the nano-sized structures of the filmare facing away from the microneedles, i.e., the structures are exposedat the surface of the composite microneedle array. The film is engagedwith the microneedle assembly and adheres to the microneedle assemblystrongly enough that the film will not delaminate from the microneedleassembly during use.

The microneedle array, including a microneedle assembly and a filmapplied thereto that includes a plurality of nanosized structures, maybe utilized in a drug delivery application. For example, a microneedlearray may be configured to be placed in fluid communication with a drugdelivery assembly. A drug delivery assembly may include a drug compoundin fluid communication with the microneedle assembly during use. In onepreferred embodiment, a microneedle assembly may be a component of atransdermal patch for drug delivery. Beneficially, the nanosizedstructures located at the surface of the microneedle assembly mayprovide efficient delivery of active agents, including large molecularweight active agents, while decreasing immune response and foreign bodyresponse to both the drug delivery device and the agents delivered bythe device.

Various aspects of the disclosure will now be described in more detail.

I. Microneedle Assembly

The microneedle assembly contains a plurality of microneedles thatextend outwardly from a support. Referring to FIG. 1, for example, oneparticular embodiment of a microneedle assembly 10 is shown thatcontains a plurality of microneedles 12 that extend from a support 20.The support 20 may be constructed from a rigid or flexible sheet ofmetal, ceramic, plastic or other material The support 20 may vary inthickness to meet the needs of the device, such as about 1000micrometers or less, in some embodiments from about 1 to about 500micrometers, and in some embodiments, from about 10 to about 200micrometers.

FIG. 2 illustrates another embodiment of a microneedle assembly 11 eachmicroneedle 14 in fluid communication with an aperture 28 that may beformed in the support 15. The aperture 28 extends through the support15. The microneedles 14 extend from the surface of the support 15.

It should be understood that the number of microneedles shown in thefigures is for illustrative purposes only. The actual number ofmicroneedles used in a microneedle assembly may, for example, range fromabout 500 to about 10,000, in some embodiments from about 2,000 to about8,000, and in some embodiments, from about 4,000 to about 6,000.

The size and shape of the microneedles may also vary as desired. Forexample, the microneedles 14 of FIG. 2 include a cylindrical portionupon which is positioned a conical portion having a tip. In alternativeembodiments, however, the microneedles 14 may have an overall pyramidalshape or an overall conical shape. Regardless, the microneedle 14typically includes a base 320 and a tip 322. As shown in FIG. 2, thebase 320 is the portion of the microneedle 318 that is proximate to thesurface of the support 15. The tip 322 of a microneedle 14 is the pointof the microneedle that is furthest from the base 320. Although the tip322 may be variously formed, it typically has a radius that is less thanor equal to about 1 micrometer. The microneedles 14 are typically of alength sufficient to penetrate the stratum corneum and pass into theepidermis, but not penetrate through the epidermis and into the dermisin applications where it is desirable to minimize pain. In certainembodiments, the microneedles have a length (from their tip 322 to theirbase 320) between about 1 micrometer and about 1 millimeter in length,for instance about 500 micrometers or less, or between about 10micrometers and about 500 micrometers, or between about 30 micrometersand abut 200 micrometers.

The size of individual needles may be optimized depending upon thedesired targeting depth, the strength requirements of the needle toavoid breakage in a particular tissue type, etc. For instance, thecross-sectional dimension of a transdermal microneedle may be betweenabout 10 nanometers (nm) and 1 millimeter (mm), or between about 1micrometer (μm) and about 200 micrometers, or between about 10micrometers and about 100 micrometers. The outer diameter may be betweenabout 10 micrometers and about 100 micrometers and the inner diameter ofa hollow needle may be between about 3 micrometers and about 80micrometers. The tip typically has a radius that is less than or equalto about 1 micrometer.

The microneedles 14 may be arranged on the substrate in a variety ofpatterns, and such patterns may be designed for a particular use. Forexample, the microneedles 14 may be spaced apart in a uniform manner,such as in a rectangular or square grid or in concentric circles. Thespacing may depend on numerous factors, including height and width ofthe microneedles 14, the characteristics of the film that will beapplied to the surface of the microneedles 14, as well as the amount andtype of substance that is intended to be moved through the microneedles14. While a variety of arrangements of microneedles is useful, aparticularly useful arrangement of microneedles 14 is a “tip-to-tip”spacing between microneedles of about 50 micrometers or more, in someembodiments about 100 to about 800 micrometers, and in some embodiments,from about 200 to about 600 micrometers.

The microneedles 14 may be formed of various substances such as, forexample, polymers, ceramics and metals. While numerous processes may beused to manufacture microneedles, a suitable production system is MEMS(Micro-Electro-Mechanical Systems) technology and microfabricationprocesses. MEMS is capable of forming micromechanical and other elementssuch as semiconductors on a single silicon substrate usingmicrofabrication processes such as etching, micromachining or otherprocesses. The support 15 may be manufactured from silicon, themicroneedles being subsequently formed by a microetching process.Micromolding techniques may also be used to form the microneedles 14 andsupport 15.

In one embodiment, the microneedles 14 may define at least one channel16 that is in fluidic communication with at least a portion of anaperture 28 of the support 15. The dimensions of the channel 16, whenpresent, may be specifically selected to induce capillary flow of a drugcompound. Capillary flow generally occurs when the adhesive forces of afluid to the walls of a channel are greater than the cohesive forcesbetween the liquid molecules. Specifically, capillary pressure isinversely proportional to the cross-sectional dimension of the channel16 and directly proportional to the surface tension of the liquid,multiplied by the cosine of the contact angle of the fluid in contactwith the material forming the channel. Thus, to facilitate capillaryflow in the patch, the cross-sectional dimension (e.g., width, diameter,etc.) of the channel 16 may be selectively controlled, with smallerdimensions generally resulting in higher capillary pressure. Forexample, in some embodiments, the cross-sectional dimension of thechannel typically ranges from about 1 micrometer to about 100micrometers, in some embodiments from about 5 micrometers to about 50micrometers, and in some embodiments, from about 10 micrometers to about30 micrometers. The dimension may be constant or it may vary as afunction of the length of the channel 16. The length of the channel mayalso vary to accommodate different volumes, flow rates, and dwell timesfor the drug compound. For example, the length of the channel may befrom about 10 micrometers to about 800 micrometers, in some embodimentsfrom about 50 micrometers to about 500 micrometers, and in someembodiments, from about 100 micrometers to about 300 micrometers. Thecross-sectional area of the channel may also vary. For example, thecross-sectional area may be from about 50 square micrometers to about1,000 square micrometers, in some embodiments from about 100 squaremicrometers to about 500 square micrometers, and in some embodiments,from about 150 square micrometers to about 350 square micrometers.Further, the aspect ratio (length/cross-sectional dimension) of thechannel may range from about 1 to about 50, in some embodiments fromabout 5 to about 40, and in some embodiments from about 10 to about 20.In cases where the cross-sectional dimension (e.g., width, diameter,etc.) and/or length vary as a function of length, the aspect ratio canbe determined from the average dimensions.

Referring to FIGS. 3-4, the illustrated microneedles 318 contain atleast one channel 330. The channel may be located in a variety ofdifferent positions, such as in the interior of the microneedle, on anexterior surface, etc. In the embodiment illustrated in FIGS. 3-4, forexample, the channel 330 is located on an exterior surface of themicroneedle 318. The cross-section of a channel may be any suitableshape. For example the channel 16 shown in FIG. 2 is substantiallyU-shaped. The channel may be arcuate or have any other configurationsuitable for moving a substance therethrough, such as, for example,V-shaped or C-shaped. Regardless, and referring again to FIGS. 3-4, apathway 326 may be formed by the channel 330 and the aperture 328, whichmeet at a junction 332 that is generally located in the plane of thesurface 316. Each microneedle 318 may deliver or extract drug compoundsthrough the skin via the pathway 326, as depicted in FIGS. 3-4. Thepathway 326 enables the compound to flow from the first surface 314through the aperture 328, the junction 332 and exit into the channel330. By enabling the compound to flow through the support and directlyinto the channel 330, more precise control over the delivery locationand the amount of substance delivered may be provided.

In certain embodiments and as shown in FIG. 3, an aperture 328 isaligned with a single channel 330 via a junction 332. Alternately and asshown in FIG. 4, a single aperture 328 may feed two or more separatechannels 330.

The channel 330 may extend from the junction 332 at the base 320 of themicroneedle to the tip 322. In other embodiments, the channel 330 maynot extend the full length of the microneedle 318 to the tip 322, asshown in FIGS. 3-4. Each microneedle 318 may include more than onechannel 330, as in the embodiment of FIG. 4. Alternate embodiments mayinclude more channels if desired. The channel 330 may be variouslypositioned on the exterior surface 324, forming a substantially linearpath from the base 320 towards the tip 322, or forming a winding orcircuitous path along the exterior surface 324. In microneedles wheretwo or more channels are present, the channels 330 may be variouslyspaced around the microneedle 318 in a symmetrical or asymmetricalmanner.

II. Nanopatterned Film

The microneedle assembly may be draped or layered with a film thatdefines a plurality of structures on a surface. FIGS. 5A and 5Bschematically illustrate the patterned (FIG. 5A) and unpatterned (FIG.5B) sides of one embodiment of a film 500 defining a first side 501 anda second side 502. A plurality of structures 505 is formed on the firstside 501 of film 500. Film 500 may have a thickness from the first side501 to the second side 502 of greater than about 2 micrometers, in oneembodiment. For instance film 500 may have a thickness between about 1micrometer and about 1 millimeter. A film 500 may have a thicknessbetween about 5 micrometers and about 200 micrometers, or between about5 micrometers and about 100 micrometers, in one embodiment.

A film 500 may include a plurality of identical structures formed on thefirst surface 501 or may include structures formed of various sizes,shapes and combinations thereof in a random or nonrandom pattern. Apredetermined pattern of structures may include a mixture of structureshaving various lengths, diameters, cross-sectional shapes, and/orspacings between the structures. For example, the structures may bespaced apart in a uniform manner, such as in a rectangular or squaregrid or in concentric circles.

A complex nanotopography may be formed on a film that may define afractal or fractal-like geometry. As utilized herein, the term “fractal”generally refers to a geometric or physical structure or pattern havinga fragmented shape at all scales of measurement between a greatest and asmallest scale such that certain mathematical or physical properties ofthe structure behave as if the dimensions of the structure are greaterthan the spatial dimensions. Mathematical or physical properties ofinterest may include, for example, the perimeter of a curve or the flowrate in a porous medium. The geometric shape of a fractal may be splitinto parts, each of which defines self-similarity. Additionally, afractal has a recursive definition and has a fine structure atarbitrarily small scales.

As utilized herein, the term “fractal-like” generally refers to ageometric or physical structure or pattern having one or more, but notall, of the characteristics of a fractal. For instance, a fractal-likestructure may include a geometric shape that includes self-similarparts, but may not include a fine structure at an arbitrarily smallscale. In another example, a fractal-like geometric shape or physicalstructure may not decrease (or increase) in scale equally betweeniterations of scale, as may a fractal, though it will increase ordecrease between recursive iterations of a geometric shape of thepattern. A fractal-like pattern may be simpler than a fractal. Forinstance, it may be regular and relatively easily described intraditional Euclidean geometric language, whereas a fractal may not.

A film 500 may include structures of the same general shape (e.g.,pillars) and the pillars may be formed to different scales ofmeasurement (e.g., nano-scale pillars as well as micro-scale pillars).Alternatively, a film may include at a surface structures that vary inboth scale size and shape or that vary only in shape while formed to thesame nano-sized scale. Structures may vary with regard to size and/orshape and may form a complex nanotopography.

At least a portion of the structures may be nanostructures formed on anano-sized scale, e.g., defining a cross-sectional dimension of lessthan about 500 nanometers, for instance less than about 400 nanometers,less than about 250 nanometers, or less than about 100 nanometers. Thecross sectional dimension may generally be greater than about 5nanometers, for instance greater than about 10 nanometers, or greaterthan about 20 nanometers. For example, the nanostructures may define across sectional dimension between about 5 nanometers and about 500nanometers, between about 20 nanometers and about 400 nanometers, orbetween about 100 nanometers and about 300 nanometers. In cases wherethe cross sectional dimension of a nanostructure varies as a function ofheight of the nanostructure, the cross sectional dimension can bedetermined as an average from the base to the tip of the nanostructures,or as the maximum cross sectional dimension of the structure, forexample the cross sectional dimension at the base of a cone-shapednanostructure.

FIG. 4 illustrates one embodiment of a complex nanotopography as may beformed on a surface. This particular pattern includes a central largepillar 100 and surrounding pillars 102, 104, of smaller dimensionsprovided in a regular pattern. As may be seen, this pattern includes aniteration of pillars, each of which is formed with the same generalshape, but vary with regard to horizontal dimension. This particularcomplex pattern is an example of a fractal-like pattern that does notinclude identical alteration in scale between successive recursiveiterations. For example, while the pillars 102 are first nanostructuresthat define a horizontal dimension that is about one third that of thelarger pillar 100, which is a microstructure, the pillars 104 are secondnanostructures that define a horizontal dimension that is about one halfthat of the pillars 102.

A pattern that includes structures of different sizes may include largerstructures having a cross-sectional dimension formed on a larger scale,e.g., microstructures having a cross-sectional dimension greater thanabout 500 nanometers in combination with smaller nanostructures. In oneembodiment, microstructures of a complex nanotopography may have across-sectional dimension between about 500 nanometers and about 10micrometers, between about 600 nanometers and about 1.5 micrometers, orbetween about 650 nanometers and about 1.2 micrometers. For example, thecomplex nanotopography of FIG. 8 includes micro-sized pillars 100 havinga cross sectional dimension of about 1.2 micrometers.

When a pattern includes one or more larger microstructures, forinstance, having a cross-sectional dimension greater than about 500nanometers, determined either as the average cross sectional dimensionof the structure or as the largest cross sectional dimension of thestructure, the complex nanotopography will also include nanostructures,e.g., first nanostructures, second nanostructures of a different sizeand/or shape, etc. For example, pillars 102 of the complexnanotopography of FIG. 6 have a cross-sectional dimension of about 400nanometers, and pillars 104 have a cross-sectional dimension of about200 nanometers.

A nanotopography may be formed of any number of different elements. Forinstance, a pattern of elements may include two different elements,three different elements, an example of which is illustrated in FIG. 6,four different elements, or more. The relative proportions of therecurrence of each different element may also vary. In one embodiment,the smallest elements of a pattern will be present in larger numbersthan the larger elements. For instance in the pattern of FIG. 6, thereare eight pillars 104 for each pillar 102, and there are eight pillars102 for the central large pillar 100. As elements increase in size,there may generally be fewer recurrences of the element in thenanotopography. By way of example, a first element that is about 0.5,for instance between about 0.3 and about 0.7 in cross-sectionaldimension as a second, larger element may be present in the topographyabout five times or more than the second element. A first element thatis approximately 0.25, or between about 0.15 and about 0.3 incross-sectional dimension as a second, larger element may be present inthe topography about 10 times or more than the second element.

The spacing of individual elements may also vary. For instance,center-to-center spacing of individual structures may be between about50 nanometers and about 1 micrometer, for instance between about 100nanometers and about 500 nanometers. For example, center-to-centerspacing between structures may be on a nano-sized scale. For instance,when considering the spacing of nano-sized structures, thecenter-to-center spacing of the structures may be less than about 500nanometers. This is not a requirement of a topography, however, andindividual structures may be farther apart. The center-to-center spacingof structures may vary depending upon the size of the structures. Forexample, the ratio of the average of the cross-sectional dimensions oftwo adjacent structures to the center-to-center spacing between thosetwo structures may be between about 1:1 (e.g., touching) and about 1:4,between about 1:1.5 and about 1:3.5, or between about 1:2 and about 1:3.For instance, the center to center spacing may be approximately doublethe average of the cross-sectional dimensions of two adjacentstructures. In one embodiment, two adjacent structures each having across-sectional dimension of about 200 nanometers may have acenter-to-center spacing of about 400 nanometers. Thus, the ratio of theaverage of the diameters to the center-to-center spacing in this case is1:2.

Structure spacing may be the same, i.e., equidistant, or may vary forstructures in a pattern. For instance, the smallest structures of apattern may be spaced apart by a first distance, and the spacing betweenthese smallest structures and a larger structure of the pattern orbetween two larger structures of the pattern may be the same ordifferent as this first distance.

For example, in the pattern of FIG. 6, the smallest structures 104 havea center-to-center spacing of about 200 nanometers. The distance betweenthe larger pillars 102 and each surrounding pillar 104 is less, about100 nanometers. The distance between the largest pillar 100 and eachsurrounding pillar 104 is also less than the center-to-center spacingbetween to smallest pillars 104, about 100 nanometers. Of course, thisis not a requirement, and all structures may be equidistant from oneanother or any variation in distances. In one embodiment, differentstructures may be in contact with one another, for instance atop oneanother, as discussed further below, or adjacent one another and incontact with one another.

Structures of a topography may all be formed to the same height,generally between about 10 nanometers and about 1 micrometer, but thisis not a requirement, and individual structures of a pattern may vary insize in one, two, or three dimensions. In one embodiment, some or all ofthe structures of a topography can have a height of less than about 20micrometers, less than about 10 micrometers, or less than about 1micrometer, for instance less than about 750 nanometers, less than about680 nanometers, or less than about 500 nanometers. For instance thestructures can have a height between about 50 nanometers and about 20micrometers or between about 100 nanometers and about 700 nanometers.For example, nanostructures or microstructures can have a height betweenabout 20 nm and about 500 nm, between about 30 nm and about 300 nm, orbetween about 100 nm and about 200 nm, though it should be understoodthat structures may be nano-sized in a cross sectional dimension and mayhave a height that may be measured on a micro-sized scale, for instancegreater than about 500 nm. Micro-sized structures can have a height thatis the same or different from nano-sized structures of the same pattern.For instance, micro-sized structures can have a height of between about500 nanometers and about 20 micrometers, or between about 1 micrometerand about 10 micrometers, in another embodiment. Micro-sized structuresmay also have a cross sectional dimension on a micro-scale greater thanabout 500 nm, and may have a height that is on a nano-sized scale ofless than about 500 nm.

The aspect ratio of the structures (the ratio of the height of astructure to the cross sectional dimension of the structure) can bebetween about 0.15 and about 30, between about 0.2 and about 5, betweenabout 0.5 and about 3.5, or between about 1 and about 2.5. For instance,nanostructures may have an aspect ratio falling within any of theseranges.

A film surface may include a single instance of a pattern, as shown inFIG. 6, or may include multiple iterations of the same or differentpatterns. For example, FIG. 7 illustrates a surface pattern includingthe pattern of FIG. 6 in multiple iterations over a surface.

FIGS. 8A and 8B illustrate increasing magnification images of anotherexample of a complex nanotopography. The nanotopography of FIGS. 8A and8B includes an array of fibrous-like pillars 70 located on a substrate.At the distal end of each individual pillar, the pillar splits intomultiple smaller fibers 60. At the distal end of each of these smallerfibers 60, each fiber splits again into multiple filaments (not visiblein FIGS. 8A and 8B). Structures formed on a surface that have an aspectratio greater than about 1 may be flexible, as are the structuresillustrated in FIGS. 8A and 8B, or may be stiff.

FIGS. 8C and 8D illustrate another example of a complex nanotopography.In this embodiment, a plurality of pillars 72 each including an annularhollow therethrough 71 are formed on a substrate. At the distal end ofeach hollow pillar, a plurality of smaller pillars 62 is formed. As maybe seen, the pillars of FIGS. 8C and 8D maintain their stiffness andupright orientation. Additionally, and in contrast to previous patterns,the smaller pillars 62 of this embodiment differ in shape from thelarger pillars 72. Specifically, the smaller pillars 62 are not hollow,but are solid. Thus, nanotopography including structures formed to adifferent scale need not have all structures formed with the same shape,and structures may vary in both size and shape from the structures of adifferent scale.

FIG. 9 illustrates another pattern including nano-sized structures asmay be formed on the surface of the film. As may be seen, in thisembodiment, individual pattern structures may be formed at the samegeneral size, but with different orientations and shapes from oneanother.

The pattern of the individual structures may be designed so as to affectthe packing density. For instance, square packing (FIG. 10A), hexagonalpacking (FIG. 10B), or some variation thereof may be utilized to patternthe structures on a film. When designing a pattern in which varioussized structures of cross sectional areas A, B, and C are adjacent toone another on a substrate, circle packing as indicated in FIG. 10C maybe utilized. Of course, variations in packing density and determinationof associated alterations in characteristics of a surface are wellwithin the abilities of one of skill in the art.

The formation of nanotopography on a film surface may increase thesurface area of the film without a corresponding increase in volume. Ingeneral, the surface area to volume ratio of the film may be greaterthan about 10,000 cm⁻¹, greater than about 150,000 cm⁻¹, or greater thanabout 750,000 cm⁻¹. Determination of the surface area to volume ratiomay be carried out according to any standard methodology as is known inthe art. For instance, the specific surface area of a surface may beobtained by the physical gas adsorption method (B.E.T. method) withnitrogen as the adsorption gas, as is generally known in the art anddescribed by Brunauer, Emmet. and Teller (J. Amer. Chem, Soc., vol. 60,February., 1938, pp. 309-319), incorporated herein by reference. The BETsurface area may be less than about 5 m²/g, in one embodiment, forinstance between about 0.1 m²/g and about 4.5 m²/g, or between about 0.5m²/g and about 3.5 m²/g. Values for surface area and volume may also beestimated from the geometry of molds used to form a surface, accordingto standard geometric calculations. For example, the volume may beestimated according to the calculated volume for each pattern elementand the total number of pattern elements in a given area, e.g., over thesurface of a single microneedle.

The nanotopography of a film may be characterized through determinationof the fractal dimension of the pattern of structures on the film. Inone embodiment, nanotopography of a surface of the device may exhibit afractal dimension of greater than about 1, for instance between about1.2 and about 5, between about 1.5 and about 3. or between about 1.5 andabout 2.5.

The fractal dimension is a statistical quantity that gives an indicationof how completely a fractal appears to fill space as the recursiveiterations continue to smaller and smaller scale. The fractal dimensionof a two dimensional structure may be represented as:

$D = \frac{\log \; {N(e)}}{\log (e)}$

where N(e) is the number of self-similar structures needed to cover thewhole object when the object is reduced by 1/e in each spatialdirection.

For example, when considering the two dimensional fractal known as theSierpenski triangle illustrated in FIG. 11, in which the mid-points ofthe three sides of an equilateral triangle are connected and theresulting inner triangle is removed, the fractal dimension is calculatedas follows:

$D = \frac{\log \; {N(e)}}{\log (e)}$$D = \frac{\log \; 3}{\log \; 2}$ D ≈ 1.585

Thus, the Sierpenski triangle fractal exhibits an increase in linelength over the initial two dimensional equilateral triangle.Additionally, this increase in line length is not accompanied by acorresponding increase in area.

The fractal dimension of the pattern illustrated in FIG. 6 isapproximately 1.84. In one embodiment, nanotopography of a surface ofthe device may exhibit a fractal dimension of greater than about 1, forinstance between about 1.2 and about 5, between about 1.5 and about 3,or between about 1.5 and about 2.5.

A film surface may be characterized by measurement of surface roughness.Methods for determining surface roughness are generally known in theart. For instance, an atomic force microscope process in contact ornon-contact mode may be utilized according to standard practice todetermine the surface roughness of a material. Surface roughness thatmay be utilized to characterize a microneedle may include the averagesurface roughness (R_(A)), the root mean square roughness, the skewness,and/or the kurtosis. In general, the average surface roughness (i.e.,the arithmetical mean height of the surface are roughness parameter asdefined in the ISO 25178 series) of a surface defining a fabricatednanotopography thereon may be less than about 200 nanometers, less thanabout 190 nanometers, less than about 100 nanometers, or less than about50 nanometers. For instance, the average surface roughness may bebetween about 10 nanometers and about 200 nanometers, or between about50 nanometers and about 190 nanometers.

The film may be characterized by the elastic modulus of thenanopatterned surface, for instance by the change in elastic modulusupon the addition of a nanotopography to a surface. In general, theaddition of a plurality of structures forming nanotopography on asurface may decrease the elastic modulus of a material, as the additionof nano-sized structures on a surface will lead to a reduction incontinuity of the surface and a related change in surface area. Ascompared to a similar surface formed according to the same process andof the same materials, but for a pattern of nanotopography on thesurface, the film including nanotopography thereon may exhibit adecrease in elastic modulus of between about 35% and about 99%, forinstance between about 50% and about 99%, or between about 75% and about80%. By way of example, the effective compression modulus of ananopatterned surface may be less than about 50 MPa, or less than about20 MPa. In one embodiment the effective compression modulus may bebetween about 0.2 MPa and about 50 MPa, between about 5 MPa and about 35MPa, or between about 10 MPa and about 20 MPa. The effective shearmodulus may be less than about 320 MPa, or less than about 220 MPa. Forinstance, the effective shear modulus may be between about 4 MPa andabout 320 MPa, or between about 50 MPa and about 250 MPa, in oneembodiment.

The film including nanotopography thereon may also exhibit an increasein surface energy as compared to a similar film that does not have asurface defining a pattern of nanotopography thereon. For instance, afilm including a nanotopography formed thereon may exhibit an increasein surface energy as compared to a similar film of the same materialsand formed according to the same methods, but for the inclusion of apattern of nanotopography on a surface. For instance, the water contactangle of a surface including a nanotopography thereon may be greaterthan about 80°, greater than about 90°, greater than about 100°, orgreater than about 110°. For example, the water contact angle of asurface may be between about 80° and about 150°, between about 90° andabout 130° or between about 100° and about 120°, in one embodiment.

The film including a plurality of nanostructures thereon may be formedaccording to any known method. A nanostructured film may be formed in asingle or multi-step process. For instance, a film may first be formed,and then a pattern of nanostructures may be molded or built onto asurface of the film. In another embodiment, a single step process inwhich the film is formed with the structures thereon may be carried out.

In one embodiment, a master defining a plurality of nanostructuresthereon may be formed and then a film may be molded through pressmolding against the master. The pattern of structures may be formed on amaster according to any nanofabrication method including, withoutlimitation, nanoimprinting, lithography, embossing molding, and soforth.

Lithography techniques, including photolithography, e-beam lithography,X-ray lithography, and so forth may be utilized for primary patterndefinition and formation of a master. Self-assembly technologiesincluding phase-separated block copolymer, polymer demixing andcolloidal lithography techniques may also be utilized in forming thenanotopography on the surface of the master.

Combinations of methods may be used, as is known. For instance,substrates patterned with colloids may be exposed to reactive ionetching (RIE, also known as dry etching) so as to refine thecharacteristics of a fabricated nanostructure such as nanopillardiameter, profile, height, pitch, and so forth. Wet etching may also beemployed to produce alternative profiles for fabricated nanostructuresinitially formed according to a different process, e.g., polymerde-mixing techniques.

Structure diameter, shape, and pitch may be controlled via selection ofappropriate materials and methods. For example, etching of metalsinitially evaporated onto colloidal-patterned substrates followed bycolloidal lift-off generally results in prism-shaped pillars. An etchingprocess may then be utilized to complete the structures as desired.Ordered non-spherical polymeric nanostructures may also be fabricatedvia temperature-controlled sintering techniques, which form a variety ofordered trigonal nanometric features in colloidal interstices followingselective dissolution of polymeric nanoparticles. These and othersuitable formation processes are generally known in the art (see, e.g.,Wood, J R Soc Interface, 2007 Feb. 22; 4(12): 1-17, incorporated hereinby reference).

Other methods as may be utilized in forming a master include utilizationof ultra-high precision laser machining techniques, examples of whichhave been described by Hunt, et al. (U.S. Pat. No. 6,995,336) and Guo,et al. (U.S. Pat. No. 7,374,864), both of which are incorporated hereinby reference.

Structures may also be formed according to chemical addition processes.For instance, film deposition, sputtering, chemical vapor deposition(CVD), epitaxy (vapor phase, liquid phase, and molecular beam),electroplating, and so forth may be utilized for building nanostructureson a master.

Self-assembled monolayer (SAM) processes as are known in the art may beutilized to form a pattern of nanostructures. For instance, the abilityof block copolymers to self-organize may be used to form a monolayerpattern on a surface, The pattern may then be used as a template for thegrowth of desired structures, e.g., colloids, according to the patternof the monolayer.

By way of example, a two-dimensional, cross-linked polymer network maybe produced from monomers with two or more reactive sites. Suchcross-linked monolayers have been made using SAM (e.g., a gold/alkylthiol system) or Langmuir-Blodgett (LB) monolayer techniques (Ahmed etal., Thin Solid Films 187: 141-153 (1990)) as are known in the art. Themonolayer may be crosslinked, which may lead to formation of a morestructurally robust monolayer.

The monomers used to form a patterned monolayer may incorporate all thestructural moieties necessary to affect the desired polymerizationtechnique and/or monolayer formation technique, as well as to influencesuch properties as overall solubility, dissociation methods, andlithographic methods. A monomer may contain at least one and more oftenat least two, reactive functional groups.

A molecule used to form an organic monolayer may include any of variousorganic functional groups interspersed with chains of methylene groups.For instance a molecule may be a long chain carbon structure containingmethylene chains to facilitate packing. The packing between methylenegroups may allow weak Van der Waals bonding to occur, enhancing thestability of the monolayer produced and counteracting the entropicpenalties associated with forming an ordered phase. In addition,different terminal moieties such as hydrogen-bonding moieties may bepresent at one terminus of the molecules, in order to allow growth ofstructures on the formed monolayer, in which case the polymerizablechemical moieties may be placed in the middle of the chain or at theopposite terminus. Any suitable molecular recognition chemistry may beused in forming the assembly. For instance, structures may be assembledon a monolayer based on electrostatic interaction, Van der Waalsinteraction, metal chelation, coordination bonding (i.e., Lewisacid/base interactions), ionic bonding, covalent bonding, or hydrogenbonding.

When utilizing a SAM-based system, an additional molecule may beutilized to form the template. This additional molecule may haveappropriate functionality at one of its termini in order to form a SAM.For example, on a gold surface, a terminal thiol may be included. Thereare a wide variety of organic molecules that may be employed to effectreplication. Topochemically polymerizable moieties, such as dienes anddiacetylenes, are particularly desirable as the polymerizing components.These may be interspersed with variable lengths of methylene linkers.

For an LB monolayer, only one monomer molecule is needed because themolecular recognition moiety may also serve as the polar functionalgroup for LB formation purposes. Lithography may be carried out on a LBmonolayer transferred to a substrate, or directly in the trough. Forexample, an LB monolayer of diacetylene monomers may be patterned by UVexposure through a mask or by electron beam patterning.

Monolayer formation may be facilitated by utilizing molecules thatundergo a topochemical polymerization in the monolayer phase. Byexposing the assembling film to a polymerization catalyst, the film maybe grown in situ, and changed from a dynamic molecular assembly to amore robust polymerized assembly for use as the master.

Techniques useful in patterning a monolayer include, but are not limitedto, photolithography, e-beam techniques, focused ion-beam techniques,and soft lithography. Various protection schemes such as photoresist maybe used for a SAM-based system. Likewise, block copolymer patterns maybe formed on gold and selectively etched to form patterns. For atwo-component system, patterning may also be achieved with readilyavailable techniques.

Soft lithography techniques may be utilized to pattern the monolayer inwhich ultraviolet light and a mask may be used for patterning. Forinstance, an unpatterned base monolayer may be used as a platform forassembly of a UV/particle beam reactive monomer monolayer. The monomermonolayer may then be patterned by UV photolithography, e-beamlithography, or ion beam lithography, even though the base SAM is notpatterned.

Growth of structures on a patterned monolayer may be achieved by variousgrowth mechanisms, such as through appropriate reduction chemistry of ametal salts and the use of seed or template-mediated nucleation. Usingthe recognition elements on the monolayer, inorganic growth may becatalyzed at this interface by a variety of methods. For instanceinorganic compounds in the form of colloids bearing the shape of thepatterned organic monolayer may be formed. For instance calciumcarbonate or silica structures may be templated by various carbonylfunctionalities such as carboxylic acids and amides. By controlling thecrystal growth conditions, it is possible to control the thickness andcrystal morphology of the mineral growth. Titanium dioxide may also betemplated.

Templated electroless plating techniques may be used to synthesizemetals using existing organic functional groups. In particular, bychelating metal atoms to the carbonyl moieties of the organic pattern,electroless metal deposition may be catalyzed on the pattern, formingpatterned metallic colloids. For instance, Cu, Au, Ni, Ag, Pd, Pt andmany other metals plateable by electroless plating conditions may beused to form metal structures in the shape of the organic monolayer. Bycontrolling the electroless plating conditions, it is possible tocontrol the thickness of the plated metal structures.

Other ‘bottom-up’ type growth methods as are known in the art may beutilized for forming the master, for example a method as described inU.S. Pat. No. 7,189,435 Tuominen, et al., which is incorporated hereinby reference, may be utilized. According to this method, a conducting orsemiconducting substrate (for example, a metal, such as gold) may becoated with a block copolymer film (for example, a block copolymer ofmethylmethacrylate and styrene), where one component of the copolymerforms nanoscopic cylinders in a matrix of another component of thecopolymer. A conducting layer may then be placed on top of the copolymerto form a composite structure. Upon vertically orientation of thecomposite structure, some of the first component may be removed, forinstance by exposure to UV radiation, an electron beam, or ozone,degradation, or the like to form nanoscopic pores in that region of thesecond component.

In another embodiment, described in U.S. Pat. No. 6,926,953 to Nealey,et al., incorporated herein by reference, copolymer structures may beformed by exposing a substrate with an imaging layer thereon, forinstance an alkylsiloxane or an octadecyltrichlorosilane self assembledmonolayer, to two or more beams of selected wavelengths to forminterference patterns at the imaging layer to change the wettability ofthe imaging layer in accordance with the interference patterns. A layerof a selected block copolymer, for instance a copolymer of polystyreneand poly(methyl methacrylate) may then be deposited onto the exposedimaging layer and annealed to separate the components of the copolymerin accordance with the pattern of wettability and to replicate thepattern of the imaging layer in the copolymer layer. Stripes or isolatedregions of the separated components may thus be formed with periodicdimensions in the range of 100 nanometers or less.

A master may be utilized to form the desired structures on a flexiblefilm. The master may be a positive master or a negative master,depending upon the molding method used to form the film from the master.For instance, the master may be a negative mold master, defining thenegatives of the nanostructures to be formed on a film, and the film maybe formed or molded against the negative master through application ofheat and/or pressure. In another embodiment, a positive master may beused to form a negative master, for instance via an eletroformingprocess, and the negative master may then be used in a film formingprocess. A process of electroforming involves placing the positive moldmaster into an electroforming tank that deposits a metal around thefeatures of the master. This may be any suitable metal. The metal isdeposited to a desired thickness at which point the positive mold masteris separated from the electroformed metal creating the negative moldcomponents. This form of a mold is typically called an electroform.Following formation, an electroform may then be cut utilized to mold afilm surface.

In one embodiment, a pattern from a master may be transferred to a filmaccording to a process and device as described in U.S. Pat. No.7,754,131 to Olsson, et al., which is incorporated herein by reference.For instance, the master may be located in conjunction with an aligned,nonpatterned film. The master may then be pressed against the film undersufficient heat and pressure so as to transfer the pattern to the film.Preferred pressures and temperatures may depend upon the material thatforms the film. For example, the film may be heated to a temperature upto about 500° C. prior to being pressed against the master. Othermethods and devices for forming a pattern of nanostructures on a film asare known in the art may alternatively be utilized such as thosedescribed in U.S. Pat. Nos. 6,923,930, 7,022,465, 7,041,228, 7,137,336,7,195,734, 7,252,492, 7,670,127, 7,670,452, 7,687,007, 7,704,425,7,717,693, 7,855,046, and 7,862,849, all of which are incorporatedherein by reference.

The film may generally be a biocompatible polymeric film formed of amoldable thermoplastic or thermoset polymer. By way of example, a filmmay include homopolymers and/or copolymers of a polyolefin (e.g.,polypropylene, polyethylene), a polystyrene, a polycarbonate, apolymethacrylate (e.g., polymethylmethacrylate), and so forth. In oneembodiment, a film may include biodegradable polymers as are known inthe art including, without limitation, homopolymers and copolymers ofpolylactides (e.g., PLA-co-PGA copolymers), polyhydroxyalkanoates, andthe like.

III. Formation Of Composite Array

The film formed with a plurality of nanosized structures on at least oneside may be applied to the microneedle assembly to form the compositemicroneedle array. FIG. 12 schematically illustrates the ends of tworepresentative microneedles 22 that include a film 26 defining afabricated nanostructure on the surface of the microneedles 22. In thisparticular embodiment, microneedles 22 define a central bore 24 as maybe used for delivery of an agent via the microneedles 22. The surface 25of microneedle 22 has been layered with a film 26 that defines a patternof nanostructures thereon. In this particular embodiment, thenanotopography of film 26 defines a random pattern on the surface.

FIGS. 13A-13C schematically illustrate a formation process for acomposite microneedle array. Microneedle assembly 418 includes amicroneedle 414 formed on a surface. In FIG. 13A, both a side view and atop view of microneedle 414 may be seen. Microneedle 414 includeschannels 412 and apertures 410 aligned with channels 412. FIG. 13Billustrates a film 420 includes a plurality of nanostructures 424 formedon a first side of the film 420. As shown in FIG. 13C, a compositemicroneedle array 430 includes the film 420 applied to the surface ofthe microneedle 414.

The film 420 is draped over and engaged with the microneedle 414 so thatit conforms to at least a portion of the shape of the microneedle. Thismay be accomplished in one embodiment through application of pressure tothe film 420, optionally in conjunction with heat to conform the film tothe shape of the microneedle as well as to adhere the film to themicroneedle surface. For example, the draped microneedle may be held ona heated vacuum box (e.g., about 3 inches H₂O vacuum) at an elevatedtemperature that is below the softening point of the polymeric materialthat forms the film 420 (e.g., between about 100 ° C. and about 130° C.)for a period of time (e.g., between about 15 minutes and about one hour)to at least partially conform the film to the surface of themicroneedles while maintaining the nanopatterned surface of the film.Thus, in this application, the film 420 is pulled down to engage withthe surface of the microneedle 414. The preferred pressure andtemperature for any specific system may depend upon the specificmaterials involved, e.g., the polymeric material of the film, as well asthe size and materials of the microneedles of the microneedle assembly.

In another embodiment, a film 420 may be engaged with the surface of themicroneedle 414 through application of pressure to the top of the film420, optionally with the application of heat. For example, the film 420may be draped over the surface of a microneedle assembly and force maybe applied to the film surface to push the film 420 against the assemblyand so engage and at least partially conform the film 420 to the surfaceof the microneedles of the assembly. In one embodiment, a negative ofthe surface of the microneedle assembly may be pressed against the film420 so as to engage the film 420 with the microneedle assembly 418. Inone embodiment, a pressing device may contact the film at certainlocations of the film, for instance where the film 420 meets the surface440 of the microneedle assembly that is between individual microneedles,and/or where the film 420 overlays the channels 412, and may avoidcontact with the film 420 where the film 420 overlays the microneedles414, so as to avoid damage to the nanostructures 424 through excessforce being placed upon the nanostructures 424 during the engagementprocess.

In one embodiment, a pressing device may apply direct pressure to thefilm 420 at those locations where the film 420 overlays the channels412. This may serve to engage the film 420 with the microneedle assembly418 and may also form a perforation in the film 420 at the channels 412,which may aid flow of an agent through the composite microneedle device.

In the embodiment of FIG. 13, the film 420 is continuous over the tip422 of the microneedle 414. During use, pressure may be applied to thedevice, for instance in applying a transdermal patch to the skin surfaceof a user, and during application a perforation may form in the film 420at or near the microneedle tip 422. According to another embodiment,pressure may be applied to the microneedle tip 422 prior to use, forinstance during engagement of the film 420 with the microneedle array418, or during formation of a drug delivery device that incorporates thecomposite microneedle array 430, and this may form a perforation in thefilm 420 that is layered over the microneedle assembly 418. Aperforation in the film 420 at the microneedle tip 422 and/or at thechannels 412 provides a route for an agent to be delivered from the drugdelivery device to a delivery site. According to another embodiment, thefilm may be formed so as to be porous to the agent(s) to be delivered bythe composite array, and the agents may diffuse across the film via theporosity, for instance upon exit from channels of the microneedles.

The surface of a composite microneedle array may be functionalized forimproved interaction with tissues or individual cells during use. Forinstance, one or more biomolecules such as polynucleotides,polypeptides, entire proteins, polysaccharides, and the like may bebound to a structured surface prior to use. In some embodiments, asurface including nanosized structures thereon may already containsuitable reactivity such that additional desired functionality mayspontaneously attach to the surface with no pretreatment of the surfacenecessary. However, in other embodiments, pretreatment of the structuredsurface prior to attachment of the desired compound may be carried out.For instance, reactivity of a structure surface may be increased throughaddition or creation of amine, carboxylic acid, hydroxy, aldehyde,thiol, or ester groups on the surface. In one representative embodiment,a microneedle surface including a pattern of nanostructures formedthereon may be aminated through contact with an amine-containingcompound such as 3-aminopropyltriethoxy silane in order to increase theamine functionality of the surface and bind one or more biomolecules tothe surface via the added amine functionality. The functionalization ofthe film surface may be carried out prior to formation of the structureson the surface. Additionally, the functionalization of the film surfacemay take place prior to applying the film to the surface of themicroneedle assembly or following the application process, as desired.

Materials as may be desirably bound to the surface of a compositemicroneedle array may include ECM proteins such as laminins,tropoelastin and/or elastin, tropocollagen and/or collagen, fibronectin,and the like. Short polypeptide fragments may be bound to the surface ofa patterned device such as an RGD sequence, which is part of therecognition sequence of integrin binding to many ECM proteins. Thus,functionalization of a nanostructured surface with RGD may encourageinteraction of the device with ECM proteins and further limit foreignbody response to the device during use.

IV. Representative Methods of Use

The composite microneedle array may be used for interaction with tissue,such as in delivery of a bioactive agent to a cell. For example, themicroneedle array may be a component of a transdermal patch used todeliver an agent to the tissue or to one or more cell types of thetissue, or may be used for structural support of a tissue, for removalof a portion or component of the tissue, and so forth. The microneedlearray may be used in one embodiment for transport of a substance acrossone or more layers of the skin.

During use, the nanotopography of the array may interact withsurrounding biological components and regulate or modulate (i.e.,change) intracellular and/or intercellular signal transductionassociated with cell/cell interactions, endocytosis, inflammatoryresponse, and so forth. For instance, through interaction between thenanotopography on a surface of the microneedles and surroundingbiological materials or structures, the device may regulate and/ormodulate membrane potential, membrane proteins, and/or intercellularjunctions (e.g., tight junctions, gap junctions, and/or desmasomes). Themicroneedle array may be utilized for transdermal delivery of agents orwithdrawal of materials across biological barriers such as the skin, theblood-brain barrier, mucosal tissues, blood and lymph vessels, and soforth without instigating a foreign body or immune response.

Structures of the nanotopography may mimic and/or interact with one ormore ECM protein such as collagen, laminin, fibronectin, etc. Cells inthe local area surrounding the microneedles may maintain ananti-inflammatory microenvironment as the microneedle surfaces maybetter mimic the local environment either directly or indirectly, e.g.,due to protein adsorption at the surface. Thus, materials may bedelivered by use of the device without development of a foreign body orimmune response.

In one embodiment, the nanotopography of an array may interact with oneor more components of the contacting epithelial tissue to increaseporosity of the tissue via paracellular and/or transcellular transportmechanisms. Epithelial tissue as may be rendered more porous by use of acomposite microneedle array may include both simple and stratifiedepithelium, including both keratinized epithelium and transitionalepithelium. Epithelial tissue encompassed herein may include any celltypes of an epithelial layer including, without limitation,keratinocytes, squamous cells, columnar cells, cuboidal cells andpseudostratified cells.

Interaction of the nanotopography of a device with components of a cellnetwork or layer of the epidermis may modulate (i.e., change) thestructure of intercellular junctions therein. An intracellular junctionmay be at least one junction selected from the group consisting of tightjunctions, gap junctions, and desmasomes. By way of example, interactionbetween biological components and structures of the nanotopography maymodulate proteins of a cellular network so as to induce the opening oftight junctions of the stratum granulosum, thereby providing improveddelivery of an active agent across the epidermis, and in one particularembodiment, a high molecular weight active agent. Tight junctions havebeen found in the stratum granulosum and opening of the tight junctionsmay provide a paracellular route for improved delivery of active agents,particularly large molecular weight active agents and/or agents thatexhibit low lipophilicity that have previously been blocked fromtransdermal delivery.

Due to improved interaction with surrounding biological components, thedevices may facilitate improved uptake of a delivered agent. Forexample, the pharmacokinetic (PK) profile (i.e., the profile ofabsorption through the epithelial membranes) of a protein therapeuticmay be enhanced through utilization of a device including a pattern ofnanotopography. By way of example, a protein therapeutic having amolecular weight of over 100 kDa, for instance between about 100 kDa andabout 200 kDa, or about 150 kDa, may be delivered transdermally via apatch including a composite microneedle array. In one embodiment, apatch may be utilized to deliver a single dose of the proteintherapeutic, for instance between about 200 and about 500 μL, or about250 μL. Following attachment of the transdermal patch to the skin, therecipient may exhibit a PK profile that reflects a rapid rise in bloodserum concentration up to between about 500 and about 1000 nanogramstherapeutic per milliliter per square centimeter of patch area, forinstance between about 750 and about 850 nanograms therapeutic permilliliter per square centimeter patch area, within about 1 to about 4hours of administration. This initial rapid rise in blood serum level,which reflects rapid uptake of the therapeutic across the dermalbarrier, may be followed by a less rapid decline of blood serumconcentration over between about 20 and about 30 hours, for instanceover about 24 hours, down to a negligible blood serum concentration ofthe therapeutic. Moreover, the rapid uptake of the delivered therapeuticmay be accompanied by little or no inflammation. Specifically, inaddition to promoting improved delivery of an agent across a transdermalbarrier, the devices may also limit foreign body response and otherundesirable reactions, such as inflammation. Use of previously knowndevices, such as transdermal patches with no nanotopography defined atthe skin contacting surface, often led to local areas of inflammationand irritation.

V. Devices

Devices may be associated with an agent for delivery via the microneedlearray. For instance, a transdermal microneedle patch may be utilized fordelivery of materials beneath the stratum corneum to the stratumspinosum or the stratum germinativum, or even deeper into the dermis. Ingeneral, an agent may be transported across the stratum corneum inconjunction with the microneedle, e.g., within the microneedle, at thesurface of the microneedle, or at the film surface that covers themicroneedle.

The device may include a reservoir, e.g., a vessel, a porous matrix,etc., that may store an agent and provide the agent for delivery. Thedevice may include a reservoir within the device itself. For instance,the device may include a hollow, or multiple pores that may carry one ormore agents for delivery. The agent may be released from the device viadegradation of a portion or the entire device or via diffusion of theagent from the device.

FIGS. 14A and 14B are perspective views of a device including areservoir. The device 110 includes a reservoir 112 defined by animpermeable backing layer 114 and a microneedle array 116. The backinglayer and the microneedle array 116 are joined together about the outerperiphery of the device, as indicated at 118. The impermeable backinglayer 114 may be joined by an adhesive, a heat seal or the like. Thedevice 110 also includes a plurality of microneedles 120. A releaseliner 122 may be removed prior to use of the device to exposemicroneedles 120.

A formulation including one or more agents may be retained within thereservoir 112. Materials suitable for use as impermeable backing layer114 may include materials such as polyesters, polyethylene,polypropylene and other synthetic polymers. The material is generallyheat or otherwise sealable to the backing layer to provide a barrier totransverse flow of reservoir contents.

Reservoir 112, defined by the space or gap between the impermeablebacking layer 114 and the base of the microneedle array 116, provides astorage structure in which to retain the suspension of agents to beadministered. The reservoir may be formed from a variety of materialsthat are compatible with an agent to be contained therein. By way ofexample, natural and synthetic polymers, metals, ceramics, semiconductormaterials, and composites thereof may form the reservoir,

In one embodiment, the reservoir may be attached to the base upon whichthe microneedles are located. According to another embodiment, thereservoir may be separate and removably connectable to the microneedlearray or in fluid communication with the microneedle array, for instancevia appropriate tubing, leur locks, etc.

The device may include one or a plurality of reservoirs for storingagents to be delivered. For instance, the device may include a singlereservoir that stores a single agent or multiple agent formulation, orthe device may include multiple reservoirs, each of which stores one ormore agents for delivery to all or a portion of the array ofmicroneedles. Multiple reservoirs may each store a different materialthat may be combined for delivery. For instance, a first reservoir maycontain an agent, e.g., a drug, and a second reservoir may contain avehicle, e.g., saline. The different agents may be mixed prior todelivery. Mixing may be triggered by any means, including, for example,mechanical disruption (i.e. puncturing, degradation, or breaking),changing the porosity, or electrochemical degradation of the walls ormembranes separating the chambers. Multiple reservoirs may containdifferent active agents for delivery that may be delivered inconjunction with one another or sequentially.

The reservoir may be in fluid communication with one or moremicroneedles of the transdermal device, and the microneedles may definea structure (e.g., a central or lateral bore) to allow transport ofdelivered agents beneath the barrier layer.

In alternative embodiments, a device may include a microneedle assemblyand a reservoir assembly with flow prevention between the two prior touse. For instance, a device may include a release member positionedadjacent to both a reservoir and a microneedle array. The release membermay be separated from the device prior to use such that during use thereservoir and the microneedle array are in fluid communication with oneanother. Separation may be accomplished through the partial or completedetachment of the release member. For example, referring to FIGS. 15-20,one embodiment of a release member is shown that is configured to bedetached from a transdermal patch to initiate the flow of a drugcompound. More particularly, FIGS. 15-16 show a transdermal patch 300that contains a drug delivery assembly 370 and a microneedle assembly380. The drug delivery assembly 370 includes a reservoir 306 positionedadjacent to a rate control membrane 308.

The rate control membrane may help slow down the flow rate of the drugcompound upon its release. Specifically, fluidic drug compounds passingfrom the drug reservoir to the microneedle assembly via microfluidicchannels may experience a drop in pressure that results in a reductionin flow rate. If this difference is too great, some backpressure may becreated that may impede the flow of the compound and potentiallyovercome the capillary pressure of the fluid through the microfluidicchannels. Thus, the use of the rate control membrane may ameliorate thisdifference in pressure and allow the drug compound to be introduced intothe microneedle at a more controlled flow rate. The particularmaterials, thickness, etc, of the rate control membrane may vary basedon multiple factors, such as the viscosity of the drug compound, thedesired delivery time, etc,

The rate control membrane may be fabricated from permeable,semi-permeable or microporous materials that are known in the art tocontrol the rate of drug compounds and having permeability to thepermeation enhancer lower than that of drug reservoir. For example, thematerial used to form the rate control membrane may have an average poresize of from about 50 nanometers to about 5 micrometers, in someembodiments from about 100 nanometers to about 2 micrometers, and insome embodiments, from about 300 nanometers to about 1 micrometer (e.g.,about 600 nanometers). Suitable membrane materials include, forinstance, fibrous webs (e.g., woven or nonwoven), apertured films,foams, sponges, etc., which are formed from polymers such aspolyethylene, polypropylene, polyvinyl acetate, ethylene n-butyl acetateand ethylene vinyl acetate copolymers. Such membrane materials are alsodescribed in more detail in U.S. Pat. Nos. 3,797,494, 4,031,894,4,201,211, 4,379,454, 4,436,741, 4,588,580, 4,615,699, 4,661,105,4,681,584, 4,698,062, 4,725,272, 4,832,953, 4,908,027, 5,004,610,5,310,559, 5,342,623, 5,344,656, 5,364,630, and 6,375,978, which areincorporated in their entirety herein by reference for all relevantpurposes. A particularly suitable membrane material is available fromLohmann Therapie-Systeme.

Referring to FIGS. 15-16, although optional, the assembly 370 alsocontains an adhesive layer 304 that is positioned adjacent to thereservoir 306. The microneedle assembly 380 likewise includes a support312 from which extends a plurality of microneedles 330 having channels331, such as described above. The layers of the drug delivery assembly370 and/or the microneedle assembly 380 may be attached together ifdesired using any known bonding technique, such as through adhesivebonding, thermal bonding, ultrasonic bonding, etc.

Regardless of the particular configuration employed, the patch 300 alsocontains a release member 310 that is positioned between the drugdelivery assembly 370 and the microneedle assembly 380. While therelease member 310 may optionally be bonded to the adjacent support 312and/or rate control membrane 308, it is typically desired that it isonly lightly bonded, if at all, so that the release member 310 may beeasily withdrawn from the patch 300. If desired, the release member 310may also contain a tab portion 371 (FIGS. 15-16) that extends at leastpartly beyond the perimeter of the patch 300 to facilitate the abilityof a user to grab onto the member and pull it in the desired direction.In its “inactive” configuration as shown in FIGS. 15-16, the drugdelivery assembly 370 of the patch 300 securely retains a drug compound307 so that it does not flow to any significant extent into themicroneedles 330. The patch may be “activated” by simply applying aforce to the release member so that it is detached from the patch.

Referring to FIGS. 17-18, one embodiment for activating the patch 300 isshown in which the release member 310 is pulled in a longitudinaldirection. The entire release member 310 may be removed as shown inFIGS. 19-20, or it may simply be partially detached as shown in FIGS.17-18. In either case, however, the seal previously formed between therelease member 310 and the aperture (not shown) of the support 312 isbroken. In this manner, a drug compound 107 may begin to flow from thedrug delivery assembly 170 and into the channels 131 of the microneedles130 via the support 112. An exemplary illustration of how the drugcompound 307 flows from the reservoir 306 and into the channels 331 isshown in FIGS. 19-20. Notably, the flow of the drug compound 307 ispassively initiated and does not require any active displacementmechanisms (e.g., pumps).

In the embodiments shown in FIGS. 15-20, the detachment of the releasemember immediately initiates the flow of the drug compound to themicroneedles because the drug delivery assembly is already disposed influid communication with the microneedle assembly. In certainembodiments, however, it may be desired to provide the user with agreater degree of control over the timing of the release of the drugcompound. This may be accomplished by using a patch configuration inwhich the microneedle assembly is not initially in fluid communicationwith the drug delivery assembly. When it is desired to use the patch,the user may physically manipulate the two separate assemblies intofluid communication. The release member may be separated either beforeor after such physical manipulation occurs.

Referring to FIGS. 21-26, for example, one particular embodiment of apatch 200 is shown. FIGS. 21-22 illustrate the patch 200 before use, andshows a first section 250 formed by a microneedle assembly 280 and asecond section 260 formed by a drug delivery assembly 270. The drugdelivery assembly 270 includes a reservoir 206 positioned adjacent to arate control membrane 208 as described above. Although optional, theassembly 270 also contains an adhesive layer 204 that is positionedadjacent to the reservoir 206. The microneedle assembly 280 likewiseincludes a support 212 from which extends a plurality of microneedles230 having channels 231, such as described above.

In this embodiment, the support 212 and the rate control membrane 208are initially positioned horizontally adjacent to each other, and arelease member 210 extends over the support 212 and the rate controlmember 208. In this particular embodiment, it is generally desired thatthe release member 210 is releasably attached to the support 212 and therate control membrane 208 with an adhesive (e.g., pressure-sensitiveadhesive). In its “inactive” configuration as shown in FIGS. 21-22, thedrug delivery assembly 270 of the patch 200 securely retains a drugcompound 207 so that it does not flow to any significant extent into themicroneedles 230. When it is desired to “activate” the patch, therelease member 210 may be peeled away and removed, such as illustratedin FIGS. 23-24, to break the seal previously formed between the releasemember 210 and the aperture (not shown) of the support 212. Thereafter,the second section 260 may be folded about a fold line “F” as shown bythe directional arrow in FIG. 25 so that the rate control member 208 ispositioned vertically adjacent to the support 212 and in fluidcommunication therewith. Alternatively, the first section 250 may befolded. Regardless, folding of the sections 250 and/or 260 initiates theflow of a drug compound 207 from the drug delivery assembly 270 and intothe channels 231 of the microneedles 230 via the support 212 (See FIG.26).

The device may deliver an agent at a rate so as to be therapeuticallyuseful. In accord with this goal, a transdermal device may include ahousing with microelectronics and other micro-machined structures tocontrol the rate of delivery either according to a preprogrammedschedule or through active interface with the patient, a healthcareprofessional, or a biosensor. The device may include a material at asurface having a predetermined degradation rate, so as to controlrelease of an agent contained within the device. A delivery rate may becontrolled by manipulating a variety of factors, including thecharacteristics of the formulation to be delivered (e.g., viscosity,electric charge, and/or chemical composition); the dimensions of eachdevice (e.g., outer diameter and the volume of any openings); the numberof microneedles on a transdermal patch; the number of individual devicesin a carrier matrix; the application of a driving force (e.g., aconcentration gradient, a voltage gradient, a pressure gradient); theuse of a valve; and so forth.

Transportation of agents through the device may be controlled ormonitored using, for example, various combinations of valves, pumps,sensors, actuators, and microprocessors. These components may beproduced using standard manufacturing or microfabrication techniques.Actuators that may be useful with the device may include micropumps,microvalves, and positioners. For instance, a microprocessor may beprogrammed to control a pump or valve, thereby controlling the rate ofdelivery.

Flow of an agent through the device may occur based on diffusion orcapillary action, or may be induced using conventional mechanical pumpsor nonmechanical driving forces, such as electroosmosis orelectrophoresis, or convection. For example, in electroosmosis,electrodes are positioned on a biological surface (e.g., the skinsurface), a microneedle, and/or a substrate adjacent a microneedle, tocreate a convective flow which carries oppositely charged ionic speciesand/or neutral molecules toward or into the delivery site.

Flow of an agent may be manipulated by selection of the material formingthe microneedle surface. For example, one or more large grooves adjacentthe microneedle surface of the device may be used to direct the passageof drug, particularly in a liquid state. Alternatively, the physicalsurface properties of the device may be manipulated to either promote orinhibit transport of material along the surface, such as by controllinghydrophilicity or hydrophobicity.

The flow of an agent may be regulated using valves or gates as is knownin the art. Valves may be repeatedly opened and closed, or they may besingle-use valves. For example, a breakable barrier or one-way gate maybe installed in the device between a reservoir and the patternedsurface. When ready to use, the barrier may be broken or gate opened topermit flow through to the microneedle surface. Other valves or gatesused in the device may be activated thermally, electrochemically,mechanically, or magnetically to selectively initiate, modulate, or stopthe flow of molecules through the device. In one embodiment, flow iscontrolled by using a rate-limiting membrane as a “valve.'”

In general, any agent delivery control system, including reservoirs,flow control systems, sensing systems, and so forth as are known in theart may be incorporated with devices. By way of example, U.S. Pat. Nos.7,250,037, 7,315,758, 7,429,258, 7,582,069, and 7,611,481 describereservoir and control systems as may be incorporated in devices.

Agents as may be delivered by the device may be intended for the localarea near the device or may be intended for wider distribution. Forinstance, in one embodiment, the device may deliver agents for painmanagement or inflammation management to a local area around a joint,for instance in treatment of osteoarthritis or rheumatoid arthritis.

The nanotopography of the device may improve delivery of agents whileminimizing foreign body and immune response. This may prove particularlybeneficial when considering delivery of oligonucleotides and othertherapeutics to the nuclear envelope. In the past, delivery of materials(e.g., plasmids, siRNA, RNAi, and so forth), to the nuclear envelope hasproven problematic because even when endocytosis is achieved, properendosomal delivery to the nuclear envelope has proven difficult, mostlikely due to foreign body and immune response. Once in the cytoplasm,delivered material is often recycled via late endosomes or degraded inthe lysosome. By use of disclosed devices, interaction of a microneedlewith the ECM may prevent foreign body response within a cell followingendocytosis and encourage delivery of the materials to the nucleus.

Delivery of protein therapeutics has likewise proven problematic in thepast. For instance, delivery of high molecular weight agents such asprotein therapeutics has proven difficult for transdermal deliveryroutes due to the natural barriers of the skin. The presence of thenanotopography on a microneedle may beneficially affect thethermodynamics of the ECM and improve efficiency of delivery and uptakeof protein therapeutics. As utilized herein, the term ‘proteintherapeutics’ generally refers to any biologically active proteinaceouscompound including, without limitation, natural, synthetic, andrecombinant compounds, fusion proteins, chimeras, and so forth, as wellas compounds including the 20 standard amino acids and/or syntheticamino acids. For instance, the presence of the device in or near thestratum granulosum may open tight junctions and allow paracellulartransport of high molecular weight agents. In one embodiment, the devicemay be utilized in transdermal delivery of high molecular weight agents(e.g., agents defining a molecular weight greater than about 400 Da,greater than about 10 kDa, greater than about 20 kDa, or greater thanabout 100 kDa, e.g., about 150 kDa). Additionally, variation of thesurface area to volume ratio of the device may be utilized to alterprotein adsorption at the surface of the device, which may in turn alterdelivery and cellular uptake of materials. Thus, deliver of a particularmaterial may be optimized through optimization of the surfacearea/volume ratio of the device.

Even when considering delivery of small molecular weight agents, thedevice may provide increased efficiency and improved uptake due tointeraction of the device with components of the dermal connectivetissue and accompanying decrease in foreign body response andimprovement in localized chemical potential of the area.

Of course, devices are not limited to targeted delivery of agents.Systemic deliver of agents is also encompassed herein as is withdrawalof an agent from a subject via the device.

There is no particular limitation to agents as may be delivered by useof the device. Agents may include proteinaceous agents such as insulin,immunoglobulins (e.g., IgG, IgM, IgA, IgE), TNF-α, antiviralmedications, and so forth; polynucleotide agents including plasmids,siRNA, RNAi, nucleoside anticancer drugs, vaccines, and so forth; andsmall molecule agents such as alkaloids, glycosides, phenols, and soforth. Agents may include anti-infection agents, hormones, drugsregulating cardiac action or blood flow, pain control, and so forth.Still other substances which may be delivered in accordance with thepresent disclosure are agents useful in the prevention, diagnosis,alleviation, treatment, or cure of disease. A non-limiting listing ofagents includes anti-Angiogenesis agents, anti-depressants, antidiabeticagents, antihistamines, anti-inflammatory agents, butorphanol,calcitonin and analogs, COX-II inhibitors, dermatological agents,dopamine agonists and antagonists, enkephalins and other opioidpeptides, epidermal growth factors, erythropoietin and analogs, folliclestimulating hormone, glucagon, growth hormone and analogs (includinggrowth hormone releasing hormone), growth hormone antagonists, heparin,hirudin and hirudin analogs such as hirulog, IgE suppressors and otherprotein inhibitors, immunosuppressives, insulin, insulinotropin andanalogs, interferons, interleukins, leutenizing hormone, leutenizinghormone releasing hormone and analogs, monoclonal or polyclonalantibodies, motion sickness preparations, muscle relaxants, narcoticanalgesics, nicotine, non-steroid anti-inflammatory agents,oligosaccharides, parathyroid hormone and analogs, parathyroid hormoneantagonists, prostaglandin antagonists, prostaglandins, scopolamine,sedatives, serotonin agonists and antagonists, sexual hypofunction,tissue plasminogen activators, tranquilizers, vaccines with or withoutcarriers/adjuvants, vasodilators, major diagnostics such as tuberculinand other hypersensitivity agents as described in U.S. Pat. No.6,569,143 entitled “Method of Intradermally Injecting Substances”, theentire content of which is incorporated herein by reference. Vaccineformulations may include an antigen or antigenic composition capable ofeliciting an immune response against a human pathogen or from otherviral pathogens.

In one preferred embodiment, the device may be utilized in treatment ofa chronic condition, such as rheumatoid arthritis, to deliver a steadyflow of an agent, to a subject in need thereof. RA drugs that may bedelivered via disclosed devices may include symptom suppressioncompounds, such as analgesics and anti-inflammatory drugs including bothsteroidal and non-steroidal anti-inflammatory drugs (NSAID), as well asdisease-modifying antirheumatic drugs (DMARDs).

The device may include and deliver symptom suppression compounds, suchas analgesics and anti-inflammatory drugs, as well as DMARD compounds,including biological DMARDs. While not wishing to be bound to anyparticular theory, it is understood that the nanometer-scale structuresfabricated on the surface of the device improve deliver of the compoundsacross the dermal barrier. Through utilization of the device, RA drugsmay be delivered at a steady concentration over a sustained period. Thedevice may prevent the initial burst of concentration common whenutilizing previously known methods for delivery of RA drugs, includingoral delivery and injection.

The present disclosure may be further understood with reference to theExamples provided below.

EXAMPLE 1

Several different molds were prepared using photolithography techniquessimilar to those employed in the design and manufacture of electricalcircuits. Individual process steps are generally known in the art andhave been described.

Initially, silicon substrates were prepared by cleaning with acetone,methanol, and isopropyl alcohol, and then coated with a 258 nanometer(nm) layer of silicon dioxide according to a chemical vapor depositionprocess.

A pattern was then formed on each substrate via an electron beamlithography patterning process as is known in the art using a JEOLJBX-9300FS EBL system. The processing conditions were as follows:

-   -   Beam current=11 nA    -   Acceleration voltage=100 kV    -   Shot pitch=14 nm    -   Dose=260 μC/cm²    -   Resist=ZEP520A, ˜330 nm thickness    -   Developer=n-amyl acetate    -   Development=2 min. immersion, followed by 30 sec. isopropyl        alcohol rinse.

A silicon dioxide etch was then carried out with an STS Advanced OxideEtch (AOE). Etch time was 50 seconds utilizing 55 standard cubiccentimeters per minute (sccm) He, 22 sccm CF₄, 20 sccm C₄F₈ at 4 mTorr,400 W coil, 200 W RIE and a DC Bias of 404-411 V.

Following, a silicon etch was carried out with an STS silicon oxide etch(SOE). Etch time was 2 minutes utilizing 20 sccm 012 and 5 sccm Ar at 5mTorr, 600 W coil, 50 W RIE and a DC Bias of 96-102 V. The silicon etchdepth was 500 nanometers.

A buffered oxide etchant (BOE) was used for remaining oxide removal thatincluded a three minute BOE immersion followed by a deionized waterrinse.

An Obducat NIL-Eitre®6 nanoimprinter was used to form nanopatterns on avariety of polymer film substrates. External water was used as coolant.The UV module utilized a single pulsed lamp at a wave length of between200 and 1000 nanometers at 1.8 W/cm². A UV filter of 250-400 nanometerswas used. The exposure area was 6 inches with a maximum temperature of200° C. and 80 Bar. The nanoimprinter included a semi-automaticseparation unit and automatic controlled demolding.

To facilitate the release of the nanoimprinted films from the molds, themolds were treated with Trideca-(1,1,2,2-tetrahydro)-octytrichlorosilane(F₁₃-TCS). To treat a mold, the silicon mold was first cleaned with awash of acetone, methanol, and isopropyl alcohol and dried with anitrogen gas. A Petri dish was placed on a hot plate in a nitrogenatmosphere and 1-5 ml of the F₁₃-TCS was added to the Petri dish. Asilicon mold was placed in the Petri dish and covered for 10-15 minutesto allow the F₁₃-TCS vapor to wet out the silicon mold prior to removalof the mold.

Five different polymers as given in Table 1, below, were utilized toform various nanotopography designs.

TABLE 1 Glass Surface Transition Tensile Tension Temperature, Modulus(mN/m) Polymer T_(g) (K.) (MPa) @20° C. Polyethylene 140-170 100-300 30Polypropylene 280 1,389 21 PMMA 322 3,100 41 Polystyrene 373 3,300 40Polycarbonate 423 2,340 43

Several different nanotopography patterns were formed, schematicrepresentations of which are illustrated in FIGS. 27A-27E. Thenanotopography pattern illustrated in FIG. 27E was a surface of a flatsubstrate purchased from NTT Advanced Technology of Tokyo, Japan. Thepatterns were designated DN1 (FIG. 27A), DN2 (FIG. 27B), DN3 (FIG. 27C),DN4 (FIG. 27D) and NTTAT2 (FIG. 27E). SEM images of the molds are shownin FIGS. 27A, 27B, and 27C, and images of the films are shown in FIGS.27D and 27E. FIG. 11 illustrates a nanopatterned film formed by use ofthe mold of FIG. 27A (DN1). In this particular film, the polymerfeatures were drawn by temperature variation as previously discussed.The surface roughness of the pattern of FIG. 27E was found to be 34nanometers.

The pattern illustrated in FIGS. 100 and 10D was also formed accordingto this nanoimprinting process. This pattern included the pillars 72 andpillars 62, as illustrated. Larger pillars 72 were formed with a 3.5micrometer (μm) diameter and 30 μm heights with center-to-center spacingof 6.8 μm. Pillars 62 were 500 nanometers in height and 200 nanometersin diameter and a center-to-center spacing of 250 nanometers.

The nanoimprinting process conditions used with polypropylene films areprovided below in Table 2.

TABLE 2 Temperature Pressure Time (s) (C.) (Bar) 10 50 10 10 75 20 10100 30 420 160 40 180 100 40 180 50 40 180 25 40

EXAMPLE 2

Films were formed as described in Example 1 including various differentpatterns and formed of either polystyrene (PS) or polypropylene (PP).The underlying substrate varied in thickness. Patterns utilized wereDN2, DN3, or DN4 utilizing formation processes as described inExample 1. The pattern molds were varied with regard to hole depth andfeature spacing to form a variety of differently-sized features havingthe designated patterns. Sample no, 8 (designated BB1) was formed by useof a 0.6 μm millipore polycarbonate filter as a mold. A 25 μmpolypropylene film was laid over the top of the filter and was thenheated to melt such that the polypropylene could flow into the pores ofthe filter. The mold was then cooled and the polycarbonate molddissolved by use of a methylene chloride solvent.

SEMs of the formed films are shown in FIGS. 28-36 and thecharacteristics of the formed films are summarized in Table 3, below.

TABLE 3 Film Cross Surface Water Sample thickness Pattern SectionalFeature Aspect Roughness Fractal Contact No. FIG. Pattern Material (μm)Feature¹ Dimension² height³ Ratio (nm) Dimension Angle 1 28 DN3 PS 75 A1100 nm 520 nm 0.47 150 2.0 100° B 400 nm 560 nm 1.4 C 200 nm 680 nm 3.42 29A, 29B DN2 PP 5.0 n/a 200 nm 100 nm 0.5 16 2.15  91° 3 30 DN2 PS 75n/a 200 nm 1.0 μm 5 64 2.2 110° 4 31 DN2 PP 25.4 n/a 200 nm 300 nm 1.538 1.94 118° 5 32 DN3 PS 75 A 1100 nm 570 nm 0.52 21.1 1.98 100° B 400nm 635 nm 1.6 C 200 nm — — 6 33 DN4 PS 75 n/a 200 nm — — 30.6 2.04  80°7 34 DN4 PP 25.4 n/a 200 nm — — 21.4 2.07 112° 8 35 BB1 PP 25.4 n/a 600nm 18 μm 30 820 2.17 110° 9 36 DN3 PP 5 A 1100 nm 165 nm 0.15 50 2.13 —B 400 nm 80 nm 0.2 C 200 nm 34 nm 6.17 ¹Pattern Features as shown on thefigures. ²Cross sectional dimension values were derived from the moldand equated as an approximation of the maximum dimension of thestructure, although it should be understood that the actual dimension ofany given individual structure may vary slightly as may be seen in thefigures. ³Feature heights are provided as the average of severalindividually determined feature heights

For each sample AFM was utilized to characterize the film.Characterizations included formation of scanning electron micrograph(SEM), determination of surface roughness, determination of maximummeasured feature height, and determination of fractal dimension.

The atomic force microscopy (AFM) probe utilized was a series 16 siliconprobe and cantilever available from μMasch. The cantilever had aresonant frequency of 170 kHz, a spring constant of 40 N/m, a length of230±5 μm, a width of 40±3 μm, and a thickness of 7.0±0.5 μm. The probetip was an n-type phosphorous-doped silicon probe, with a typical probetip radius of 10 nanometers, a full tip cone angle of 40°, a total tipheight of 20-25 μm, and a bulk resistivity 0.01-0.05 ohm-cm.

The surface roughness value given in Table 3 is the arithmetical meanheight of the surface areal roughness parameter as defined in the ISO25178 series.

The Fractal Dimension was calculated for the different angles byanalyzing the Fourier amplitude spectrum; for different angles theamplitude Fourier profile was extracted and the logarithm of thefrequency and amplitude coordinates calculated. The fractal dimension,D, for each direction is then calculated as

D=(6+s)/2,

where s is the (negative) slope of the log-log curves. The reportedfractal dimension is the average for all directions.

The fractal dimension may also be evaluated from 2D Fourier spectra byapplication of the Log Log function. If the surface is fractal the LogLog graph should be highly linear, with at negative slope (see, e.g.,Fractal Surfaces, John C. Russ, Springer-Verlag New York, LLC, July,2008).

EXAMPLE 3

An array of microneedles including an overlaid film defining ananopatterned surface was formed. Initially, an array of microneedleswas formed on a silicon wafer via a photolithography process. Eachneedle included two oppositely placed side channels, aligned with onethrough-die hole in the base of the needle.

Microneedles were formed according to a typical micromachining processon a silicon based wafer. The wafers were layered with resist and/oroxide layers followed by selective etching (oxide etching, DRIE etching,iso etching), resist stripping, oxide stripping, and lithographytechniques (e.g., iso lithography, hole lithography, slit lithography)according to standard methods to form the array of microneedles.

Following formation of the microneedle array, a 5 μm polypropylene filmincluding a DN2 pattern formed thereon as described in Example 1, thecharacteristics of which are described at sample 2 in Table 3, was laidover the microneedle array. The wafer/film structure was held on aheated vacuum box (3 inches H₂O vacuum) at elevated temperature (130°C.) for a period of one hour to gently pull the film over the surface ofthe microneedles while maintaining the nanopatterned surface of thefilm.

FIG. 37 illustrates the film over the top of the array of microneedles,and FIG. 38 is a closer view of a single needle of the array includingthe nanopatterned film overlaying the top of the needle.

EXAMPLE 4

Transdermal patches including microneedle arrays formed as described inExample 3 were formed. Patches were formed with either a DN2 pattern ora DN3 pattern on the microneedle array.

The films defining the patterns that were applied to the microneedlesare described in Table 4, below. Film 1 is equivalent to sample no. 2 ofTable 3 and Film 2 is equivalent to sample no. 9 of Table 3.

TABLE 4 Property Film 1 Film 2 Pattern DN2 DN3 Material polypropylenepolypropylene Film Thickness  5 micrometers  5 micrometers Height ofstructures 100 nm 165 nm, 80 nm, 34 nm Aspect ratio of 0.5 0.18structures Average Surface  16 nm 50 nm Roughness R_(A) FractalDimension 2.15 2.13

Control patches were also formed that had no pattern formed on the filmand subsequently applied to the array of microneedles. Transdermal andsubcutaneous formulations of etanercept (Enbrel®) were preparedaccording to instructions from the drug supplier. The subcutaneous doseformulation (for the positive control) was prepared to facilitate a4mg/kg subcutaneous drug dose. The concentration of Enbrel® fortransdermal delivery was adjusted such that an intended dosing of 200mg/kg was achieved in a 24 hr period.

A total of 10 BALB/C mice (assigned designations #1-#10) were used inthe study, 8 were transdermally dosed with Enbrel® (group 1) and 2 weresubcutaneously dosed with Enbrel® (group 2) as described in Table 5,below. The transdermal patches were applied to shaved skin areas andholes formed near the microneedle tips upon application of the patch tothe skin.

TABLE 5 Blood Group Test Dose Dose Collection Animal No. Article DrugDose Route Level volume Time Points Number 1 Transdermal Enbrel ®Transdermal 5 mg/ 0.2 ml Pre-patch #1, #5 patch subject 0.5 h #2, #6 2 h#3, #7 6 h #4, #8 24 h #2, #6 72 h #3, #7 2 subcutaneous Enbrel ®Subcutaneous 4 mg/kg 0.1 ml 24 h #9, #10 delivery

Transdermal patches used included both those defining a nanotopographyon the surface (DN2 and DN3 patterns, as described above), as well aspatches with no pattern of nanotopography.

Samples of whole blood were collected at the time points indicated inTable 5. Approximately 100 to 200 μl of blood was taken via mandibularbleeding and then centrifuged at approximately 1300 rpm for 10 minutesin a refrigerated centrifuge (set at 4° C.). The resulting serum wasaspirated and transferred within 30 minutes of bloodcollection/centrifugation to appropriately labeled tubes. The tubes werefrozen and stored in the dark at ≤−70° C. until they were analyzed forlevels of Enbrel® using Human sINF-receptor ELISA kit (R&D Systems cat#DRT200). The space time between two blood samplings on the same subjectwas 24 hours, to prevent unnecessary stress placed on the subject.

FIG. 39 graphically illustrates the average PK profile of thetransdermal patches that defined nanotopography thereon. An average ofthe results for all nanotopography-including patches was used torepresent the overall effect of incorporating nanotopography inconjunction with a microneedle transdermal patch. As may be seen, theblood serum level rose rapidly to over 800 ng/mL/cm² of patch areawithin the first two hours of attachment. Following, the blood serumlevel gradually declined to negligible within 24 hours of attachment.The data used to develop FIG. 39 is provided below in Table 6.

TABLE 6 Blood serum Time (hr) concentration (ng/ml) 0 0 0.5 192.1 2249.25 6 24.4 24 7.2 65 4.0875

FIG. 40A and 40B illustrate electron microscopy cross sectional views ofthe skin that was held in contact with the patches. The images weretaken after the patches were removed (72 hours post-attachment). Thesample of FIG. 40A was in contact with a patch including ananotopography on the surface. Specifically, a DN2 pattern, as describedabove, was formed on the surface of the patch. The sample of FIG. 40Bwas held in contact with a transdermal patch that did not define apattern of nanotopography on the surface. As may be seen, the sample ofFIG. 40B shows signs of inflammation and a high density of macrophagepresence.

EXAMPLE 5

Transdermal patches including microneedle arrays formed as described inExample 3 were formed. Patches were formed with either a HN2 pattern ora DN3 pattern on the microneedle array as described in Table 4 ofExample 4. Control patches were also formed that had no pattern formedon the film subsequently applied to the array of microneedles.Transdermal and subcutaneous formulations of etanercept (Enbrel®) wereprepared according to instructions from the drug supplier.

Test subjects (rabbits) were transdermally dosed with Enbrel® or weresubcutaneously (SubQ) dosed with Enbrel®. Results are illustratedgraphically in FIG. 41, which provides the blood serum concentration inpg/ml as a function of time. The data used to develop FIG. 41 isprovided below in Table 7, below.

TABLE 7 No structure Time microneedle Subcutaneous DN3 Subcutaneous DN30 0.00 0.00 0.00 0.00 0.00 0.5 0.00 157.49 0.00 1611.21 0.00 2 0.003029.07 0.00 3504.92 497.17 6 0.00 3545.14 338.23 3699.24 796.64 12 0.003577.13 731.22 3571.80 1080.60 24 116.78 3778.71 785.49 3464.70 1924.2448 134.23 3416.73 638.18 3885.31 1006.95 72 88.68 3356.64 572.77 3803.421172.67

While the subject matter has been described in detail with respect tothe specific embodiments thereof, it will be appreciated that thoseskilled in the art, upon attaining an understanding of the foregoing,may readily conceive of alterations to, variations of, and equivalentsto these embodiments. Accordingly, the scope of the present disclosureshould be assessed as that of the appended claims and any equivalentsthereto.

1. A composite microneedle array comprising a microneedle assembly that includes a plurality of microneedles; and a film overlaying at least one microneedle of the microneedle assembly, the film having a first surface and a second surface, wherein the first surface of the film is adhered to the microneedle and at least partially conforms thereto, the film comprising a plurality of nanostructures that are arranged in a predetermined pattern on the second surface of the film.
 2. The composite microneedle array according to claim 1, at least one of the microneedles contains a channel along a length of the microneedle.
 3. The composite microneedle array according to claim 1, at least a portion of the nanostructures have a cross-sectional dimension of less than about 500 nanometers and greater than about 5 nanometers.
 4. The composite microneedle array according to claim 1, wherein the pattern further comprises microstructures, the nanostructures having a cross-sectional dimension smaller than the microstructures.
 5. The composite microneedle array according to claim 4, further comprising second nanostructures having a cross-sectional dimension less than the cross-sectional dimension of the microstructures and greater than the cross-sectional dimension of the first nanostructures.
 6. The composite microneedle array according to claim 1, wherein at least a portion of the nanostructures have a height of from about 10 nanometers to about 20 micrometers.
 7. The composite microneedle array according to claim 1, wherein at least a portion of the nanostructures have an aspect ratio of from about 0.15 to about
 30. 8. A transdermal patch comprising a the composite microneedle array, which comprises a microneedle assembly that includes a plurality of microneedles; and a film overlaying at least one microneedle of the microneedle assembly, the film having a first surface and a second surface, wherein the first surface of the film is adhered to the microneedle and at least partially conforms thereto, the film comprising a plurality of nanostructures that are arranged in a predetermined pattern on the second surface of the film.
 9. The transdermal patch according to claim 8, further comprising a reservoir for holding a drug compound.
 10. The transdermal patch according to claim 9, further comprising a rate control membrane in fluid communication with the reservoir.
 11. The transdermal patch according to claim 10, further comprising a release member that is generally impermeable to the drug compound and positioned adjacent to the rate control membrane.
 12. The transdermal patch according to claim 9, wherein the drug compound has a molecular weight of greater than about 100 kDa.
 13. A method for forming a composite microneedle array comprising: laying a film over a microneedle assembly, the microneedle assembly including a plurality of microneedles, the film having a first surface and a second surface, the film at least partially conforming to the microneedles, wherein the first surface of the film contacts the microneedle assembly, the second surface of the film comprising a plurality of nanostructures, the nanostructures being arranged in a predetermined pattern; and engaging the film with the microneedle assembly such that the film at least partially conforms to the microneedle array and adheres to the microneedle assembly.
 14. The method according to claim 13, further comprising forming the structures on the film.
 15. The method according to claim 14, wherein the film and the nanostructures are fabricated simultaneously.
 16. The method according to claim 14, wherein the structures are fabricated on the film according to a nanoimprint lithography process.
 17. The method according to claim 13, wherein the step of engaging the film with the microneedle assembly comprises the application of at least one of heat and pressure to the film.
 18. The method according to claim 17, wherein the pressure is a vacuum pressure.
 19. The method according to claim 17, wherein the pressure is applied to the second surface of the film.
 20. The method according to claim 13, further comprising perforating the film at or near the microneedle tips. 